Iron is an essential nutrient for virtually every organism, yet it can also be a potent cellular toxin. Dysregulated iron metabolism and iron overload are features of a growing number of human diseases. Some genes involved in cellular iron uptake and export have been identified, yet very little is known about inter- and intracellular iron transport, intracellular iron utilization, and the regulation of these processes. A combination of genetic, biochemical, and cell biological approaches is needed to understand iron metabolism and the role of iron in human disease. These approaches can be combined in the simple eukaryote, Saccharomyces cerevisiae. Studies of metal metabolism in budding yeast have yielded important insights into iron, copper, and zinc metabolism in both humans and pathogenic microorganisms. Genetic studies of iron metabolism in a simple eukaryote will allow us to discover new genes involved in iron homeostasis as well as to determine the cellular response to iron overload and iron deprivation. We have used a variety of strategies to identify genes that are involved in eukaryotic iron homeostasis. Using available genome and protein databases, we have grouped these newly identified genes into families and have begun their functional evaluation. We have designed a genetic screen to identify human genes that alter iron homeostasis when expressed in yeast. Human genes for the iron-storage protein ferritin have been expressed in yeast, and although the proteins are stable and assemble into the proper 24-subunit oligomer, only a small amount of iron is deposited within the ferritin core. We hypothesize that cytosolic iron chaperones that facilitate iron loading into ferritin may be expressed in human cells, but not in yeast, which do not synthesize ferritin. We have isolated multiple human cDNAs which, when expressed in a yeast cell that contains ferritin, increase the amount of iron stored in ferritin. Further characterization of these cDNAs is underway.